PB1787: GADD45A IS OVER-EXPRESSED IN THE BONE MARROW OF PATIENTS WITH DE NOVO ACUTE MYELOID LEUKEMIA REFRACTORY TO INDUCTION CHEMOTHERAPY
Authors: A. Bouchla, G. Deliconstantinos, T. Loupis, F. Kontsioti, V. Mpakou, M. Delatola, M. Pagoni, K. Theodorakopoulos, S. Papageorgiou, V. Pappa
Journal: HemaSphere (2022)

Background: Acute myeloid leukemia (AML) is a life-threatening malignant myeloid disorder with poor prognosis. Response to induction treatment determines long-term prognosis; however, mechanisms underlying response have not been thoroughly investigated. DNA damage and repair mechanisms influence not only the genetic predisposition to leukemia but are also very important for refractoriness to treatment. Aims: The aim of this study was to investigate the possible alterations in the gene expression profile of DNA damage signaling pathways in two leukemic cell lines following their exposure to chemotherapeutic agents, namely, idarubicin and cytarabine, and verify the findings in AML patient samples at diagnosis before the onset of chemotherapy. Methods: Cell lines Kasumi-1 with the t(8;21) and MV4-11 (biphenotypic B-myelomonocytic leukemia) were exposed either to idarubicin (0.1μΜ) for 6h or cytarabine (1μΜ) for 48h. Dead cells were eliminated from drug-treated cells using an appropriate commercial kit. Gene expression profiling through PCR arrays analysis (RT2 Profiler, Qiagen) was performed in triplicate after RNA extraction from untreated, chemotherapy-treated and live cells following chemotherapy exposure. Human DNA damage signaling pathway-related gene expression was evaluated and analyzed through the RT2 Profiler PCR Array data analysis tool. Moreover, mononuclear cells were isolated from bone marrow of lymphoma patients, used as controls, and from AML patients at diagnosis who subsequently received chemotherapy with the 7 + 3 regiment. Response to first-line chemotherapy was recorded. Total RNA was isolated (RNeasy plus Mini Kit, Qiagen) and quantified and cDNA was synthesized. Relative expression of genes of interest was quantified using SYBR® Green-based real-time RT-PCR (QuantiTect® Primer Assay, Qiagen). Statistics were performed using linear regression. Results: The following genes were over two-fold up-regulated in live cells from leukemic cell lines after treatment with either agent: PPP1R15A and CDKN1A in both cell lines, GADD45A and GADD45G in live MV4-11 cells and EXO1 in live Kasumi cells. These genes were selected for relative quantification; 25 control samples and 38 AML samples (28 responders and 10 non-responders to treatment) were evaluated. GADD45A in AML compared to controls was found 2-fold up-regulated in treatment responders (ns) and 9-fold up-regulated in non-responders [p=0.01]. In addition, GADD45A was overexpressed in non-responders compared to responders [p=0.02]. PPP1R15A in AML compared to controls was 12-fold up-regulated in responders [p=0.001] and 16-fold up-regulated in non-responders [p=0.001]. (Figure 1) Image:Summary/Conclusion: GADD45A controls the balance between apoptosis and autophagy via the p38 and JNK/MAPK pathways and PPP1R15A participates in growth arrest and apoptosis in response to DNA damage by attenuating the translational elongation of key transcription factors. Both genes are likely to play an important role in the pathogenesis of AML. Furthermore, GADD45A overexpression appears to be related with resistance to induction treatment, which may have significant prognostic and therapeutic implications.